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G-protein-coupled receptors (GPCRs) are the largest family of transmembrane signaling proteins in the human genome. Events in the GPCR signaling cascade have been well characterized, but the receptor composition and its membrane distribution are still generally unknown. Although there is evidence that some members of the GPCR superfamily exist as(More)
Muscle force is generated by myosin crossbridges interacting with actin. As estimated from stiffness and equatorial X-ray diffraction of muscle and muscle fibres, most myosin crossbridges are attached to actin during isometric contraction, but a much smaller fraction is bound stereospecifically. To determine the fraction of crossbridges contributing to(More)
Recent developments in light microscopy enable individual fluorophores to be observed in aqueous conditions. Biological molecules, labeled with a single fluorophore, can be localized as isolated spots of light when viewed by optical microscopy. Total internal reflection fluorescence microscopy greatly reduces background fluorescence and allows single(More)
Pleckstrin homology (PH) domains act to target proteins to the plasma membrane and intracellular vesicles by binding to specific phosphoinositol phospholipids. We have investigated the binding kinetics of PH domains found in the tail region of the molecular motor, myosin X. Using total internal reflection fluorescence microscopy, we observed binding and(More)
Over the past 10 years, advances in laser and detector technologies have enabled single fluorophores to be visualized in aqueous solution. Here, we describe methods based on total internal reflection fluorescence microscopy (TIRFM) that we have developed to study the behavior of individual protein molecules within living mammalian cells. We have used(More)
G protein-coupled receptors (GPCRs), including dopamine receptors, represent a group of important pharmacological targets. An increased formation of dopamine receptor D2 homodimers has been suggested to be associated with the pathophysiology of schizophrenia. Selective labeling and ligand-induced modulation of dimerization may therefore allow the(More)
We show prolonged contraction of permeabilized muscle fibers of the frog during which structural order, as judged from low-angle x-ray diffraction, was preserved by means of partial cross-linking of the fibers using the zero-length cross-linker 1-ethyl-3-[3-dimethylamino)propyl]carbodiimide. Ten to twenty percent of the myosin cross-bridges were(More)
DNA helicases are motor proteins that catalyze the unwinding of double-stranded DNA into single-stranded DNA using the free energy from ATP hydrolysis. Single molecule approaches enable us to address detailed mechanistic questions about how such enzymes move processively along DNA. Here, an optical method has been developed to follow the unwinding of(More)
M2 muscarinic acetylcholine receptors modulate cardiac rhythm via regulation of the inward potassium current. To increase our understanding of M2 receptor physiology we used Total Internal Reflection Fluorescence Microscopy to visualize individual receptors at the plasma membrane of transformed CHO(M2) cells, a cardiac cell line (HL-1), primary(More)
BACKGROUND Weibel-Palade bodies (WPB) are endothelial cell (EC) specific secretory organelles containing Von Willebrand factor (VWF). The temperature-dependence of Ca(2+)-driven WPB exocytosis is not known, although indirect evidence suggests that WPB exocytosis may occur at very low temperatures. Here we quantitatively analyse the temperature-dependence of(More)